Over-expression of alpha-actinin with a GFP fusion protein is sufficient to increase whole-cell stiffness in human osteoblasts.
Ann Biomed Eng
; 36(10): 1605-14, 2008 Oct.
Article
em En
| MEDLINE
| ID: mdl-18636329
ABSTRACT
Osteoblasts respond to shear stress by simultaneously increasing their whole-cell stiffness and up-regulating the cytoskeletal crosslinking protein alpha-actinin. The stiffness of reconstituted cytoskeletal networks increases following the addition of alpha-actinin, but the effect of alpha-actinin on whole-cell mechanical behavior has not been investigated. The hypothesis of this study was that increasing alpha-actinin in the cytoskeleton would be sufficient to increase whole-cell stiffness. hFOB osteoblasts were transfected with a plasmid for GFP-tagged alpha-actinin, resulting in a 150% increase in the amount of alpha-actinin. The GFP-alpha-actinin fusion protein co-fractionated with the cytoskeleton and co-localized to the same regions of the cytoskeleton as endogenous alpha-actinin. Whole-cell mechanical behavior was measured by atomic force microscopy using a 25 mum diameter microsphere as an indenter. The whole-cell stiffness of cells over-expressing GFP-alpha-actinin was 60% higher than cells expressing only endogenous alpha-actinin (p < 0.002), which was within the range of mechanical behavior observed in osteoblastic cells exposed to 1 and 2 Pa of fluid shear. These results indicate that the up-regulation of alpha-actinin synthesis in osteoblasts is sufficient to alter the whole-cell mechanical behavior and highlights the potential role of alpha-actinin to reinforce cells against mechanical loads.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Osteoblastos
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Citoesqueleto
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Actinina
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Proteínas Recombinantes de Fusão
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Proteínas de Fluorescência Verde
Limite:
Humans
Idioma:
En
Ano de publicação:
2008
Tipo de documento:
Article